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1.
Separate Assays Specific for Ascorbate Peroxidase and Guaiacol Peroxidase and for the Chloroplastic and Cytosolic Isozymes of Ascorbate Peroxidase in Plants 总被引:35,自引:0,他引:35
Ascorbate (AsA) peroxidase can be inactivated both by p-chloromercuribenzoateand by the depletion of AsA but guaiacol peroxidases, such ashorseradish peroxidase, cannot. The cytosolic isozymes of AsAperoxidase are less sensitive to depletion of AsA than the chloroplasticisozymes, which include stromal [Chen and Asada (1989) PlantCell Physiol. 30: 987] and thyla-koid-bound [Miyake and Asada(1992) Plant Cell Physiol. 33: 541] enzymes. Exploring theseproperties, we established simple methods for separate assaysof AsA peroxidase and guaiacol peroxidase and of the three isozymesof AsA peroxidase in plant extracts. These methods were usedto characterize the guaiacol peroxidases and isozymes of AsAperoxidase in plants and algae. (Received October 20, 1993; Accepted February 7, 1994) 相似文献
2.
The photodynamic activity of Neutral Red and the new monobrominated Neutral Red was studied in suspensions of Staphylococcus aureus. The effect of mannitol and sodium azide in the presence of 25 μm photosensitizer on lethal photosensitization were investigated. The results of the mechanistic evaluation of Neutral Red showed that both mannitol and sodium azide produced a completed protective effect after irradiation without significant differences between them. The evaluation of monobrominated Neutral Red also showed a protective effect of microorganisms with the addition of mannitol. Although sodium azide produced a protective effect of the photoinactivation, it was incomplete and less than that exhibited by mannitol. The results indicate that the starting reagent, Neutral Red, is a producer of radical species, acting through a type I mechanism, whereas the halogenated derivative of Neutral Red produced reactive oxygen species and a contribution of singlet molecular oxygen cannot be discarded in the photoinactivation of Staphylococcus aureus cells. These results, analyzed together with the previously evaluated properties of the dyes, allow us to explain the differences observed in the photoinactivation of Staphylococcus aureus mediated by both azine photosensitizers. 相似文献
3.
Kitajima S Ueda M Sano S Miyake C Kohchi T Tomizawa K Shigeoka S Yokota A 《Bioscience, biotechnology, and biochemistry》2002,66(11):2367-2375
Depletion of the electron donor ascorbate causes rapid inactivation of chloroplastic ascorbate peroxidase (APX) of higher plants, while cytosolic APX is stable under such conditions. Here we report the cloning of cDNA from Galdieria partita, a unicellular red alga, encoding a novel type of APX (APX-B). The electrophoretic mobility, Km values, kcat and absorption spectra of recombinant APX-B produced in Escherichia coli were measured. Recombinant APX-B remained active for at least 180 min after depletion of ascorbate. The amino-terminal half of APX-B, which forms the distal pocket of the active site, was richer in amino acid residues conserved in chloroplastic APXs of higher plants rather than cytosolic APXs. In contrast, the sequence of the carboxyl-terminal half, which forms the proximal pocket, was similar to that of the cytosolic isoform. The stability of APX-B might be due to its cytosolic isoform-like structure of the carboxyl-terminal half. 相似文献
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Fryer MJ Ball L Oxborough K Karpinski S Mullineaux PM Baker NR 《The Plant journal : for cell and molecular biology》2003,33(4):691-705
In Arabidopsis leaves, high light stress induces rapid expression of a gene encoding a cytosolic ascorbate peroxidase (APX2), whose expression is restricted to bundle sheath cells of the vascular tissue. Imaging of chlorophyll fluorescence and the production of reactive oxygen species (ROS) indicated that APX2 expression followed a localised increase in hydrogen peroxide (H2O2) resulting from photosynthetic electron transport in the bundle sheath cells. Furthermore, leaf transpiration rate also increased prior to APX2 expression, suggesting that water status may also be involved in the signalling pathway. Abscisic acid stimulated APX2 expression. Exposure of ABA-insensitive mutants (abi1-1, abi2-1) to excess light resulted in reduced levels of APX2 expression and confirmed a role for ABA in the signalling pathway. ABA appears to augment the role of H2O2 in initiating APX2 expression. This regulation of APX2 may reflect a functional organisation of the leaf to resolve two conflicting physiological requirements of protecting the sites of primary photosynthesis from ROS and, at the same time, stimulating ROS accumulation to signal responses to changes in the light environment. 相似文献
6.
The production of reactive oxygen species on the plant-cell surface is considered. Along with the plasmalemmal redox systems, cell-wall peroxidase is involved in the production of superoxide and hydrogen peroxide. Under stress conditions, some soluble peroxidase isoforms are easily secreted into the apoplast. Various membranotropic compounds, salicylic acid in particular, can also induce this process. Mobile peroxidase forms are supposed to induce the plant defense response. 相似文献
7.
The red fluorescent protein KillerRed, engineered from the hydrozoan chromoprotein anm2CP, has been reported to induce strong cytotoxicity through the chromophore assisted light inactivation (CALI) effect. Here, we present the X-ray structures of KillerRed in its native and bleached states. A long water-filled channel is revealed, connecting the methylene bridge of the chromophore to the solvent. This channel facilitates the transit of oxygen and of reactive oxygen species (ROS) formed by reaction with the excited chromophore. The functional roles of key mutations used to produce KillerRed are discussed, strong chromophore distortions in the bleached state are revealed, and mechanisms for ROS production and self protection are proposed. The presence of a partially mature, photo-resistant, green-emitting state is characterized, which accounts for enhanced CALI by “pre-bleached” KillerRed. 相似文献
8.
盐地碱蓬(Suaeda salsa)APX 基因的克隆及盐胁迫下的表达 总被引:13,自引:0,他引:13
从盐地碱蓬 (Suaedasalsa)中克隆了抗坏血酸过氧化物酶 (ascorbateperoxidase ,APX)的全长cDNA(SsAPX) ,基因注册号为AY0 34 893。SsAPX全长 1.1kb ,推导的氨基酸序列长为 2 5 0个氨基酸残基。BLAST同源性分析表明 ,该cDNA与已报告的菠菜(Spinaciaoleracea)细胞质抗坏血酸过氧化物酶基因同源性最高 ,在核苷酸水平上一致性为 87% ,在氨基酸水平上一致性为 89%。Southern杂交表明APX基因在盐地碱蓬基因组中只有 1个拷贝。盐 (NaCl 40 0mmol/L)处理不同时间后的Northern杂交分析表明盐地碱蓬中SsAPX基因在盐胁迫下表达量增加 ,而且在盐胁迫下抗坏血酸过氧化物酶的活性也显著地增加 ,说明该基因受盐诱导。推测抗坏血酸过氧化物酶可能在保护盐地碱蓬免受氧化损伤的过程中起到一定作用 相似文献
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The complete cDNA encoding human thiol-specific antioxidant protein (PRP) was isolated from a human brain cDNA library in the λZap expression vector. An open reading frame (ORF) was identified and found to encode a polypeptide of 197 aa with a Mr of 21 729. The cDNA contained 98 bp of 5′-untranslated sequence (UTR) and 259 bp of 3′-UTR containing a poly(A) signal, AATAAA. Expression of the human PRP cDNA in Escherichia coli yielded a functionally active protein. The observed local sequence homologies between human PRP and other homologous proteins whose functions have not yet been defined give important insight into elucidating the biochemical function of a new protein family which has highly conserved regions containing cysteine. 相似文献
11.
Farrukh A. Shamsi Seema Husain S. M. Hadi 《Journal of biochemical and molecular toxicology》1996,11(2):67-71
Uric acid is present in human plasma in relatively high concentrations and is considered to be a natural physiological antioxidant. We have earlier shown that in the presence of Cu(II) and molecular oxygen, uric acid causes strand breakage in DNA. In this article, we show that uric acid fluorescence is quenched by addition of DNA, indicating the formation of uric acid-DNA complex. Uric acid-Cu(II)-mediated DNA strand scission is capable of bacteriophage inactivation and such inactivation is mediated through reduction of Cu(II) to Cu(I) and the generation of oxygen-derived radicals. It is indicated that the DNA breakage is repaired in E. coli and involves the repair of DNA polymerase. © 1996 John Wiley & Sons, Inc. 相似文献
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The doxorubicin-resistant, acute myelogenous leukemia cell line, AML-2/DX100, characterized by the over-expression of multidrug resistance protein (MRP) and the down-regulation of catalase, has advantages for the screening of MRP inhibitors as well as for cytotoxic substances producing potential reactive oxygen species. The screening power of AML-2/DX100 cells for an MRP inhibitor, probenecid, was approximately 4-fold stronger than that of another resistant cell line, HL-60/Adr, over-expressing MRP. AML-2/DX100 was approximately 2- to 5-fold more sensitive to pro-oxidants such as Paraquat, H2O2 and t-butyl hydroperoxide, when compared with its parental cells. 相似文献
13.
采用Nortnern印迹法和鲁米诺化学发光法分析抗性的、近等位基因的敏感番茄细胞株阴离子过氧化物酶基因的转录和细胞反应性氧变化,发现病原真菌的激发子和外源H2O2均能促进抗性番茄细胞中阴离子过氧化物酶转录。激发子还能刺激抗性细胞中反应性氧水平暂时急剧升高。敏感番茄细胞对激发子或H2O2没有响应。Ca2 和磷脂酶C的抑制剂硫酸新霉素分别促进和抑制激发子诱导下抗性细胞反应性氧增加。 相似文献
14.
James P. Kehrer 《Free radical research》1989,5(6):305-314
The phenomenon of reperfusion injury remains poorly defined. Questions remain about whether injury occurs in addition to that produced by hypoxia or ischemia. or whether the observed changes simply reflect the unmasking of an underlying injury. Various pathological processes which occur upon the return of oxygen to hypoxic and ischemic heart tissue have been quantitated to assess the extent of reperfusion injury. yet it is not known if they reflect identical or different processes. In addition. the mechanism(s) responsible for these diverse changes may not be the same in the various model systems used to study reperfusion injury. Although reactive oxygen species clearly are formed at reperfusion. conclusive evidence that they are producing injury. particularly during the first seconds. is not available. Several sources of these reactive oxygen species have been proposed but none have been clearly linked with injury in several species or model systems. As research in the field of reperfusion injury continues. it is imperative for scientists to clearly define the system they are using so that studies examining mechanisms of cell lysis at reperfusion are not confused with those assessing the occurrence and mechanisms of damage in addition to that produced by oxygen deprivation. 相似文献
15.
《Free radical research》2013,47(6):305-314
The phenomenon of reperfusion injury remains poorly defined. Questions remain about whether injury occurs in addition to that produced by hypoxia or ischemia. or whether the observed changes simply reflect the unmasking of an underlying injury. Various pathological processes which occur upon the return of oxygen to hypoxic and ischemic heart tissue have been quantitated to assess the extent of reperfusion injury. yet it is not known if they reflect identical or different processes. In addition. the mechanism(s) responsible for these diverse changes may not be the same in the various model systems used to study reperfusion injury. Although reactive oxygen species clearly are formed at reperfusion. conclusive evidence that they are producing injury. particularly during the first seconds. is not available. Several sources of these reactive oxygen species have been proposed but none have been clearly linked with injury in several species or model systems. As research in the field of reperfusion injury continues. it is imperative for scientists to clearly define the system they are using so that studies examining mechanisms of cell lysis at reperfusion are not confused with those assessing the occurrence and mechanisms of damage in addition to that produced by oxygen deprivation. 相似文献
16.
Isolation of ergosterol peroxide and its reversion to ergosterol in the pathogenic fungus Sporothrix schenckii 总被引:2,自引:0,他引:2
da Graça Sgarbi Diana Bridon da Silva Antonio Jorge Ribeiro Carlos Iracilda Zeppone Silva Célio Lopes Angluster Jayme Alviano Celuta Sales 《Mycopathologia》1997,139(1):9-14
Ergosterol peroxide, a presumed product of the H_2O_2-dependent enzymatic oxidation of ergosterol, has been isolated from yeast forms of the pathogenic fungus Sporothrix schenckii.
The substance, which may have a role in fungal virulence, has been characterized mainly using spectroscopic methods (1H and 13C nuclear magnetic resonance and high resolution mass spectra). The purified compound showed a molecular formula of C_28H_44O_3, displaying characteristic features of epidioxy sterols and was reverted to ergosterol when submitted to S. schenckii enzymatic
extract.
This revised version was published online in June 2006 with corrections to the Cover Date. 相似文献
17.
Yutong Liu Ling Zhang Zitai Ma Li Tian Yingchi Liu Yuqing Liu 《Free radical research》2019,53(7):758-767
Chemotherapy is the most effective strategy for the treatment of metastatic breast cancer. However, P-glycoprotein (P-gp)-mediated multidrug resistance severely limits the efficacy of chemotherapy and is a major cause of the failure during chemotherapeutic treatment. In this study, we investigated the anticancer effects of combining chemotherapeutic drugs with ascorbate (AA) in human breast cancer cells. We found that combined administration of AA can improve the sensitivity of both MCF-7 and doxorubicin (Dox)-resistant MCF-7/Adr cells to Dox in vitro and in vivo by a reactive oxygen species (ROS)-dependent mechanism. Further studies proved that AA can promote the accumulation of Dox in MCF-7/Adr cells when combined with doxorubicin. AA had no effect on the expression of P-gp at the mRNA and protein levels, but could decrease its activity as demonstrated by an obvious inhibition of efflux of P-gp substrate Rh 123. AA reduced ATP levels in both MCF-7 and MCF-7/Adr cells, and pretreating AA-stimulating cells with catalase completely rescued ATP levels. With ATP reduction, we observed an increased cellular calcium and the appearance of vacuoles and micropores on the cell surface, indicating the increased cell membrane permeability in AA-treated MCF-7/Adr cells. The above results suggest that AA could promote the cellular accumulation of doxorubicin by inducing ROS-dependent ATP depletion. Clinically, a combination of AA with doxorubicin would be a novel strategy for reversal of the multidrug resistance in human breast cancer cells during chemotherapy. 相似文献
18.
AN Run DONG ChenFang LEI YanJun HAN Lu LI Ping CHEN JianMing WANG GuiRong SHI Qi GAO Chen JIANG HuiYing ZHOU Wei HAN Jun CHU YongLie DONG XiaoPing School of Medicine Xi’an JiaoTong University Xi’an China State Key Laboratory for Infectious Disease Prevention Control National Institute for Viral Disease Control Prevention Chinese Center for Disease Control Prevention YingXin Rd Beijing China 《中国科学:生命科学英文版》2008,51(7):630-639
One of the physiological functions of cellular prion protein(PrP C )is believed to work as a cellular resistance to oxidative stress,in which the octarepeats region within PrP plays an important role.However,the detailed mechanism is less clear.In this study,the expressing plasmids of wild-type PrP (PrP-PG5)and various PrP mutants containing 0(PrP-PG0),9(PrP-PG9)and 12(PrP-PG12)octarepeats were generated and PrP proteins were expressed both in E.coli and in mammalian cells.Protein aggregation and formation of carbonyl groups were clearly seen in the recombinant PrPs expressed from E.coli after treatment of H2O2.MTT and trypan blue staining assays revealed that the cells expressing the mutated PrPs within octarepeats are less viable than the cells expressing wild-type PrP.Statistically significant high levels of intracellular free radicals and low levels of glutathione peroxidase were observed in the cells transfected with plasmids containing deleted or inserted octarepeats.Remarkably more productions of carbonyl groups were detected in the cells expressing PrPs with deleted and inserted octarepeats after exposing to H2O2.Furthermore,cells expressing wild-type PrP showed stronger resistant activity to the challenge of H2O2 at certain extent than the mutated PrPs and mock. These data provided the evidences that the octarepeats number within PrP is critical for maintaining its activity of antioxidation.Loss of its protective function against oxidative stress may be one of the possible pathways for the mutated PrPs to involve in the pathogenesis of familial Creutzfeldt-Jacob diseases. 相似文献
19.
Adenosine 3',5'-pyrophosphate, previously isolated from the red alga Porphyra perforata , possesses biological properties similar to those of a cyclic AMP-like compound isolated from higher plants. Both compounds are potent inhibitors of bovine cyclic AMP-dependent protein kinase and bovine phosphodiesterase. 相似文献
20.
Catalase-deficient strains of the human pathogenic yeast Candida albicans were constructed using the URA-blaster method. The disruptant was viable and grew normally in an ordinary culture condition, but became extremely sensitive to treatment with hydrogen peroxide. No catalase activity was observed in a catalase (CCT)-gene-disrupted strain, 1F5-4-1, suggesting that there were no other catalase or catalase-like enzymes in this yeast. The disruptant was shown to be sensitive to higher temperature and to low concentrations of SDS, NP-40, or Triton X-100. After a wild-type CCT gene was reintroduced into the disruptant, catalase activity was restored and the strain became moderately sensitive to treatment with hydrogen peroxide. However, neither the temperature sensitivity nor the susceptibility to SDS observed in the disruptant was restored in the CCT-reintroduced strain. A model infection experiment using wild-type and dCCT strains showed that the disruptants disappeared more rapidly than the wild-type strain in mouse liver, lung, and spleen. These results suggest that the catalase plays a significant role in survival in the host immune system and thus leads this organism to establish infection in the host. 相似文献